KR20210002690A - Medical use - Google Patents

Abstract

The present invention relates generally to antiviral agents for use in preventing herpes virus reactivation in a patient, wherein the patient has received a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells. Further, the present invention relates to NK cells and/or NK-like T cells for use in treating malignant diseases in a patient, wherein the use is to patients receiving NK cells and/or NK-like T cell therapeutics. And administering an antiviral agent. The invention also relates to pharmaceutical compositions and kits.

Description

Medical use

The present invention relates generally to antiviral agents for use in preventing herpes virus reactivation in a patient, wherein the patient has received a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells. Further, the present invention relates to NK cells and/or NK-like T cells for use in treating malignant diseases in a patient, wherein the use is to patients receiving NK cells and/or NK-like T cell therapeutics. And administering an antiviral agent. The invention also relates to pharmaceutical compositions and kits.

Multiple myeloma (MM) is a malignant neoplasm characterized by clonal proliferation of plasma cells in the bone marrow. MM is still thought to be incurable due to the persistence of minimal residual disease (MRD), potentially due to MM cells remaining after treatment (Alici E, Bjorkstrand B, Treschow A, Aints A, Smith CI, Gahrton G, Dilber MS. Long-term follow-up of gene-marked CD34+ cells after autologous stem cell transplantation for multiple myeloma. Cancer Gene Ther. 2007;14(3):227-32]).

The use of cellular immunotherapy for cancer has been investigated since the introduction of lymphokine-activated killer (LAK) cells in the mid-1980s (Grimm EA. et al., 1982; Rosenberg S., 1985). ]). Adoptive transfer of cytotoxic effector cells with tumor cell death potential to induce a graft versus tumor effect is an attractive approach to cancer. Natural killer (NK) and NK-like T cells contribute to a relatively high cytotoxic capacity among other effector-cell populations with potential anti-tumor effects (3). However, the low percentage of these cells in peripheral blood mononuclear cells (PBMCs) and effector-cell populations, such as LAK cells, represents a barrier to their use in clinical trials and as therapeutic agents for cancer.

Our previous studies have shown that long-term ex vivo expansion and activation of autologous NK cells from MM patients can provide significantly superior cytotoxic activity against autologous tumor cells when compared to short-term activated autologous NK cells. (Alici E, Sutlu T, Bjorkstrand B, Gilljam M, Stellan B, Nahi H, et al. Autologous antitumor activity by NK cells expanded from myeloma patients using GMP-compliant components. Blood. 2008;111(6) :3155-62] and [Sutlu T, Stellan B, Gilljam M, Quezada HC, Nahi H, Gahrton G, et al. Clinical-grade, large-scale, feeder-free expansion of highly active human natural killer cells for adoptive immunotherapy using an automated bioreactor.Cytotherapy. 2010;12(8):1044-55]). The inventors also reported an effective NK cell-based treatment of MM development in animal models (Alici E, Konstantinidis KV, Sutlu T, Aints A, Gahrton G, Ljunggren HG, et al. Anti-myeloma activity of endogenous and adoptively transferred activated natural killer cells in experimental multiple myeloma model.Exp Hematol. 2007;35(12):1839-46]). The present inventors began the first phase I/II clinical trials in humans while developing a procedure for NK cell expansion in a closed automated bioreactor using ingredients that comply with clinical grade good manufacturing practice (GMP). It has been approved by the Swedish Medicinal Products Agency (EudraCT: 2010-022330-83) and the Ethics Committee (EPN: 2013/490-32) (Sutlu T, Stellan B, Gilljam M, Quezada HC , Nahi H, Gahrton G, et al. Clinical-grade, large-scale, feeder-free expansion of highly active human natural killer cells for adoptive immunotherapy using an automated bioreactor. Cytotherapy. 2010;12(8):1044-55] And Sutlu T, Alici E. Ex vivo expansion of natural killer cells: a question of function. Cytotherapy. 2011).

Very surprisingly, during clinical trials, we observed reactivation of herpes virus (especially varicella zoster virus (VZV) expressed as shingles) in patients receiving NK cell and/or NK-like T cell therapy. I did. Shingles and other conditions caused by herpes virus reactivation and/or infection are well known, unpleasant conditions, and quality of life for patients who are already seriously ill in this situation and are receiving NK cell and/or NK-like T cell therapy. Greatly reduces.

Our surprising discovery suggests a new approach for using and managing NK cell and/or NK-like T cell therapeutics.

Thus, in a first aspect, the present invention provides an antiviral agent for use in preventing herpes virus reactivation in a patient, wherein the patient comprises natural killer (NK) cells and/or NK-like T cells. Receiving a therapeutic agent, uses include administering an antiviral agent to a patient receiving NK cell and/or NK-like T cell therapy.

In a second aspect, the present invention provides the use of an antiviral agent in the manufacture of a medicament for preventing herpes virus reactivation in a patient, wherein the patient comprises natural killer (NK) cells and/or NK-like T cells. Upon receiving a therapeutic agent, an antiviral agent is administered to a patient receiving NK cell and/or NK-like T cell therapy.

In a third aspect, the present invention provides a method of preventing herpes virus reactivation in a patient, wherein the patient is receiving a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, the method comprising NK cells And/or administering an antiviral agent to a patient receiving NK-like T cell therapy.

By “herpes virus” we include a large class of DNA viruses known as Herpesviridae (or herpesvirus). Herpes simplex viruses 1 and 2 (HSV-1 and HSV-2) (also known as human herpesvirus 1 (HHV-1) and HHV2), varicella zoster virus (VZV or HHV-3), which infect humans Epstein Barr virus (EBV or HHV-4), human cytomegalovirus (HCMV or HHV-5), human herpesvirus 6A and 6B (HHV-6A and HHV-6B), human herpesvirus 7 (HHV-7) and Kaposi There are 9 known types of herpesviruses of sarcoma-associated herpesvirus (also known as KSHV, HHV-8). In total, there are more than 130 herpes viruses (Whitley RJ. Herpesviruses. In: Baron S, editor. Medical Microbiology. 4th edition. Galveston (TX): University of Texas Medical Branch at Galveston; 1996. Chapter 68)) .

At least five species of herpes viruses: HSV-1 and HSV-2 (both can cause cleft lip and genital herpes), Varicella zoster virus (causing chickenpox and shingles), Epstein Barr virus (monocytosis) And several diseases including several cancers) and cytomegaloviruses are extremely widespread among humans. More than 90% of adults are infected with at least one of these, and the dormant form of the virus remains in most people.

The term "Varicella zoster virus (VZV)" is used to describe the virus that causes varicella (chicken pox) and herpes zoster (shingles). Varicella results from the primary infection by the virus; Herpes zosters result from secondary invasion by reactivation of the same virus or infection, which in many cases can dormant for several years.

As described above and in the accompanying examples, the inventors have surprisingly confirmed reactivation of VZV in patients receiving therapeutic agents comprising natural killer (NK) cells and/or NK-like T cells. VZV is a double-stranded DNA virus, which is morphologically identical to the herpes simplex virus. It is the causative agent of both chickenpox and herpes zoster (shingles), which is accompanied by affected sensory nerves and is characterized by an inflammatory response of the dorsal nerve root and ganglion. Chickenpox develops after initial viral exposure, is a relatively mild, self-limited childhood disease, usually accompanied by a characteristic rash, but can be disseminated in immunocompromised children. Even when the clinical symptoms of chickenpox are resolved, VZV remains dominant in the immune system of infected people in the trigeminal and dorsal root ganglia (also called viral latency).

VZV reactivation in old age produces a disease known as herpes zoster or shingles. Severe shingles complications include post-herpetic neuralgia (PHN), multiple shingles, myelitis, herpes ophthalmic, or shingles free. A common shingles complication is post-shingles neuralgia (PHN), a chronic, usually exacerbating pain condition that can persist over months or even years. The risk of PHN in patients with shingles is 10% to 18%.

Pain and paresthesia are generally the first symptoms of VZV infection. Diagnosis can be difficult until the characteristic vesicular rash breaks out. The prognostic period in which symptoms can change is common. Pain, itchiness and paresthesia are common symptoms.

Patients may experience pain, helplessness and depression and/or influenza-like symptoms during acute illness. The most common presentation is shingles vesicular rash, most commonly affecting the thoracic skin segment, and erythema spots and papules develop within 24 hours and progress to blisters after a precursor disease of pain and perceptual abnormalities. The blister eventually hardens and disappears. Pain and loss of sensation are common symptoms, movement disorders also occur, and often disappear upon examination. In severe cases, actual short paralysis due to VZV slow plexus neuritis has been reported (Prevention of herpes zoster: recommendations of the Advisory Committee on Immunization Practices (ACIP). MMWR Recomm Rep. 2008;57(05):1-30). ).

The term “herpes simplex virus” (HSV) is used to describe the viruses that cause herpes simplex infection, HSV1 and HSV2.

HSV1 and HSV2 have about 50% genomic homology, but share most of the other features. Signs of herpes simplex virus infection include stomatitis, genital herpes, herpetic keratitis and dermal live human hands. Neonatal herpes simplex virus infection and herpes simplex virus encephalitis also occur.

The virus initially replicates in epithelial cells and produces characteristic blisters at the erythematous base. It then goes up the sensory nerve to the dorsal root ganglion, where it establishes a dormant after the initial period of replication. The virus spreads distally from the ganglion during reactivated infection, initiating new skin and/or mucosal lesions. HSV1 transmission is primarily oral, and herpes simplex virus 2 is primarily genital. Transmission requires intimate contact (Whitley RJ. Herpesviruses. In: Baron S, editor. Medical Microbiology. 4th edition. Galveston (TX): University of Texas Medical Branch at Galveston; 1996. Chapter 68).

The term “Epstein Barr virus (EBV)” is used to describe a herpesvirus found in cell culture of Burkitt's lymphoma. EBV is a causative agent in infectious mononucleosis and a number of other related conditions/diseases including EBV-associated lymphoma. Epstein Barr virus causes traditional mononucleosis. In immunocompromised hosts, the virus causes lymphoproliferative syndrome. In some classes, the Epstein Barr virus causes Duncan's syndrome.

Epstein Barr virus replicates in the epithelial cells and β lymphocytes of the oropharynx. Epstein Barr virus is transmitted by intimate contact, particularly through saliva exchange (Whitley RJ. Herpesviruses. In: Baron S, editor. Medical Microbiology. 4th edition. Galveston (TX): University of Texas Medical Branch at Galveston; 1996 Chapter 68]).

The term “cytomegalovirus (CMV)” is generally used to describe infections that are not conspicuous in healthy people, but can be life-threatening in immunocompromised individuals, such as those with HIV infection, organ transplant recipients, or newborns. This is found in a significant proportion of the population. Seropositivity increases with age as in EBV. Ganciclovir, which inhibits the replication of all human herpes viruses, is commonly used to treat CMV, especially retinitis. Foscannet is also licensed in the United States. Cytomegalovirus causes three clinical syndromes: (1) Congenital cytomegalovirus infection (when symptomatic) causes hepatomegalovirus, retinitis, rash and central nervous system involvement; (2) In about 10% of adolescents and adults, primary cytomegalovirus infection causes mononucleosis syndrome with fever, sickness, atypical lymphocytosis and pharyngitis; (3) Immunocompromised hosts (transplant recipients and human immunodeficiency virus [HIV] infected individuals) can develop life-threatening disseminated diseases including the lungs, gastrointestinal tract, liver, retina and central nervous system.

Cytomegaloviruses replicate primarily in the salivary glands and kidneys, and are released from saliva and urine. Replication is slow, and the virus induces characteristic giant cells with intranuclear inclusion. It is spread through close contact with infected secretions. Among the world's most prevalent viral infections is cytomegalovirus infection (Whitley RJ. Herpesviruses. In: Baron S, editor. Medical Microbiology. 4th edition. Galveston (TX): University of Texas Medical Branch at Galveston; 1996. Chapter 68]).

As is well known, infection by the virus begins when viral particles come into contact with cells with a specific type of receptor molecule on the cell surface. After the viral envelope glycoprotein binds to the cell membrane receptor, the virions are internalized and degraded, allowing the viral DNA to migrate to the cell nucleus. The replication of viral DNA and transcription of viral genes occurs within the nucleus. Infected cells transcribe lytic viral genes during symptomatic infection.

The herpes virus is known to exist latent in the host, where it can exist for many years without any obvious signs of infection. In these cells, a small number of viral genes called latency associated transcripts (LATs) accumulate. During this latent tolerogenic cycle, the virus may be asymptomatic (or dormant) in the ganglion adjacent the spinal cord (referred to as the dorsal root ganglion) and/or in the trigeminal ganglion in the cranium. Viruses are human (latent and/or inactive) and can exist in the cell (and thus the host) indefinitely. Although primary infection is usually accompanied by a limited period of time in the clinical patient, long-term latency is asymptomatic.

By “herpes virus reactivation” we can refer to the meaning of non-primary infections, such as latent and/or human and/or inactive and/or reactivation of endogenous herpes virus in patients that can cause herpes virus infection. Include. This may include the development of a condition or disease (eg, shingles) associated with a latent herpes virus infection in a patient. The present inventors described varicella zoster virus (VZV); Herpes simplex virus (HSV) types 1 and 2; Epstein Barr virus (EBV); And a herpes virus selected from the group containing cytomegalovirus (CMV). In a preferred embodiment, the herpes virus is VZV.

Reactivation of the latent virus is implicated in a number of diseases (eg, Herpes zoster and Pityriasis Rosea). Transcription of viral genes transfers from the latent associated LAT to multiple lytic genes after reactivation, which enhances replication and viral production. Usually, lytic activation causes cell death. Clinically, lytic activation is usually accompanied by the occurrence of non-specific symptoms, such as low-grade fever, headache, sore throat, sickness and rash, and clinical signs such as swollen lymph nodes with tender lymph nodes.

By “patient” we encompass the meaning of a subject who is or is intended to receive medical treatment and/or prophylaxis and/or prophylaxis of herpes virus reactivation in a subject in need thereof. The patient may be a vertebrate, such as a vertebrate mammal, for example a human or non-human mammal, such as a domestic animal (eg, cat, dog, rabbit, cow, sheep, pig, mouse or other rodent). Preferably, the patient is human.

By “antiviral agent” we include any synthetic or natural molecule or compound capable of preventing the reactivation of the herpes virus in a patient. Such antiviral agents, for example, inactivate extracellular viral particles, prevent viral attachment and/or cell entry, and/or prevent replication of the viral genome, and/or inhibit the synthesis of certain viral protein(s). Antiviral effects can be exerted by preventing and/or preventing the assembly and/or release of new infectious virions. Examples of known antiviral agents include their triphosphate forms and nucleoside analogs after phosphorylation with phosphonoformic acid and phosphonoacetic acid and analogs thereof.

It will be understood that antiviral agents are for use in the prevention of herpes virus reactivation in patients.

In one embodiment, the antiviral agent is a vaccine.

In an alternative embodiment, the antiviral agent is not a vaccine.

By “prevention of herpes virus reactivation”, the present inventors can completely or partially prevent and/or inhibit reactivation of herpes virus infection and/or conditions or diseases associated with latent herpes virus infection (eg, shingles) in a patient. And/or reducing. Prevention of reactivation is the prevention of reactivation of herpes as human in nervous tissue and/or prevention of the occurrence of symptoms in infected patients and/or viral reactivation in patients, or conditions or diseases caused by viral reactivation. And a reduction in the severity or frequency of symptoms.

In one embodiment, the patient is susceptible to herpes virus reactivation. In a further embodiment, the patient is susceptible to the development of shingles.

If the herpes virus reactivation is completely prevented, the patient will be asymptomatic for viral infection. In some embodiments, the antiviral agent can eradicate a portion of the latent virus reservoir to reduce the proportion of reactivable virus, thus preventing and/or inhibiting and/or reducing herpes virus reactivation. If herpes virus reactivation is reduced and/or inhibited, this is a clinical sign (e.g., headache, burning of the skin in the affected area, tingling, numbness or itching, generally feeling sick, high temperature (heat), and itching). It can shorten the duration of the rash that can occur as a blister.

By “NK cell and/or NK-like T cell therapeutics” we include administering NK cells and/or NK-like T cells to a patient for therapeutic purposes. The patient may have a malignant disease such as a hematologic cancer, a solid tumor or a chronic viral infection, or may be another patient in need of such treatment.

It will be understood that the term “NK cell and/or NK-like T cell therapeutic agent” refers to a therapeutic agent comprising a therapeutically effective amount of NK cells and/or NK-like T cells.

NK cells and/or NK-like T cell therapeutics will be understood as a form of adoptive cell transfer (ACT), i.e., transfer of cells to a patient, and the two terms may be used interchangeably herein. In a preferred embodiment, the cells are derived from the patient (autologous). In an alternative embodiment, the cell is from another individual (heterologous). The terms “NK cell and/or NK-like T cell therapeutic agent” and “CellProtect” may be used interchangeably herein. The protocol for making CellProtect is described below and shown in Figure 5.

In a preferred embodiment, the NK cells and/or NK-like T cells are expanded and activated ex vivo and administered to a patient in need thereof. The preparation of NK cells and/or NK-like T cells is described in prior international publication WO 2010/110734, incorporated herein by reference.

In one embodiment, the NK cell and/or NK-like T cell therapeutic agent comprises at least 10% of NK cells having the phenotype CD3 ^- CD56 ⁺ . In one embodiment, at least 30% of the NK cells are activated NK cells.

NK cells and/or NK-like T cells can be administered by infusion to reach the central nervous system (CNS), for example, via a central-venous catheter or intravenously (IV) or as cerebrospinal fluid. It can be administered intratumorally (i.e., injected directly into the tumor), for example intratumoral administration.

In a preferred embodiment, the patient is not immunodepleted. By "the patient is not immunodepleted" we include the meaning of a patient who is not immunodepleted. Immunodepletion is a non-selective method of depleting (ie, eliminating) lymphocytes, such as T cells, such as regulatory T cells. Immunodepletion can be achieved by any means known in the art, including systemic irradiation, chemotherapy, or as a result of disease processes such as leukemia or HIV/AIDS. Alternatively, immunodepletion can be achieved by administering an antibody that specifically binds to lymphocytes. Lymphopenia and immunodepletion are used interchangeably to describe the condition of decreasing lymphocyte count. In certain embodiments, the patient is immunodepleted if the number of lymphocytes in the patient decreases by at least 50%, such as at least 60%, 70%, 80% or 90% after administration of the immunodepleting agent.

By “administering an antiviral agent to a patient receiving an NK cell and/or NK-like T cell therapeutic agent”, the inventors of the present invention provide for the patient to receive the NK cell and/or NK-like T cell therapy before, at the same time as or after receiving It includes administering an antiviral agent. In one embodiment, the antiviral agent is administered to the patient concurrently with the NK cell and/or NK-like T cell therapeutic agent.

In a preferred embodiment, the NK cell and/or NK-like T cell therapeutic agent induces and/or increases herpes virus infection.

By “NK cell and/or NK-like T cell therapeutic agent induces and/or increases herpes virus reactivation”, the inventors of the present invention believe that NK cell and/or NK-like T cell therapeutic agents completely or partially regenerate herpes virus in patients. It includes the meaning of the cause of the activation, or the cause of an increase in the reactivation of the herpes virus completely or partially in the patient. For example, NK cells and/or NK-like T cell therapeutic agents induce at least 5-fold, or at least 10-fold, or at least 50-fold greater than herpes virus reactivation in patients who have not received NK cells and/or NK-like T cell therapeutics. And/or can increase.

Methods for measuring viral reactivation are well known in the art and include measuring viral copy number. In addition, the quantification of antibodies to herpes virus is commonly used as an indirect measure of herpes virus reactivation. In addition, clinical symptoms can be used to indicate herpes virus reactivation.

As described by the accompanying examples, during clinical trials, we observed reactivation of VZV and signs of shingles in a number of patients who received NK cell and/or NK-like T cell therapy. Without wishing to be bound by theory, the inventors believe that activated NK cells and/or NK-like T cells attack reservoir cells for herpes virus upon adoptive cell transmigration, eventually leading to viral reactivation due to induced stress. I think there is.

Thus, in one embodiment, the herpes virus is human (latent) in the dorsal root ganglion.

In one embodiment of the invention, the patient has a malignant disease. By “malignant disease” we include, by way of non-limiting example, diseases including cancer that progresses quickly and generally causes threatening or death within a short period of time. The present inventors have described malignant conditions such as solid tumors, viral cancers and colorectal cancer; Brain cancer (eg, medulloblastoma and glioblastoma); Neuroblastoma; Bone cancer; Neoplasms derived from epithelial cells (epithelial carcinoma); Basal cell carcinoma; Adenocarcinoma; Gastrointestinal cancer; Cleft lip cancer, oral cancer, esophageal cancer, small intestine cancer; Stomach cancer; Colon cancer; Liver cancer; Bladder cancer; Pancreatic cancer; Ovarian cancer; Cervical cancer; Lung cancer; Breast cancer; Skin cancer (eg, melanoma), squamous cell and basal cell cancer; Prostate cancer, renal cell carcinoma and sarcoma (eg, soft tissue sarcoma) include the meaning of cancer selected from the group consisting of.

In one embodiment of the present invention, the NK cell and/or NK-like T cell therapeutic agent is for use in the treatment of a malignant disease.

In a preferred embodiment, the malignant disease is a hematological cancer. By “hemologic cancer” we include types of cancer that affect the blood, bone marrow and lymph nodes, such as those selected from the group consisting of myeloma, lymphoma, leukemia and chronic myeloproliferative diseases.

In a preferred embodiment of the present invention, the hematological cancer is selected from the group consisting of myeloma, lymphoma, leukemia and/or chronic myeloproliferative disease.

In a preferred embodiment, the hematologic cancer is multiple myeloma (MM).

In one embodiment, the NK cells have the phenotype CD3 ^- CD56 ⁺ and/or the NK-like T cells have the phenotype CD3 ⁺ CD56 ⁺ .

Preferably, NK cells and NK-like T cells are expanded ex vivo. For example, expansion may occur in a closed expansion system, such as a cell culture bag in an automated bioreactor system (see Examples 1 and 5). Less preferably, NK cells and NK-like T cells are expanded in tissue culture flasks. The present inventors described this method in International Publication No. WO 2010/110734 (see “1: Ex vivo expansion of NK cells and NK-like T cells from peripheral blood”) and in Alici E, et al., Blood. 2008;111(6):3155-62].

Preferably, the expansion is performed until the total number of cells each expands at least about 10 times or until at least about 50% of the expanded cell population comprises activated NK cells and NK-like T cells. For example, at least about 50% of the expanded cell population comprises NK cells with the phenotype CD3 ^- CD56 ⁺ .

In one embodiment, NK and NK-like T cells are activated ex vivo and become cytotoxic. In one embodiment, NK cells and NK-like T cells are expanded and activated simultaneously ex vivo. By “activated” we include the meaning that NK cells and/or NK-like T cells receive an activation signal. Activated NK cells are capable of killing certain target cells deficient in MHC class I expression. NK cells must receive activation signals that can occur in various forms, of which cytokines, Fc-receptors or other activating receptors are the most important. Cells can also be activated to produce cytokines and chemokines.

Activated NK cells and NK-like T cells show increased cytotoxicity as determined by cytotoxicity in vitro. One skilled in the art can determine cytotoxicity using methods known in the art.

One way to determine if cells exhibit increased cytotoxicity is to use an in vitro assay of cell mediated cytotoxicity against K562 cells using a standard 4 hour ⁵¹ Cr-release assay. Briefly, chromium-51 was incubated with human cancer cells at 37° C. for 1 hour, washed 3 times, and simultaneously with NK cells in three E:T ratios (20:1, 6.66:1, 2.22:1). Are cultured. 5% Triton-X 100 was used to achieve total cell lysis. The supernatant was harvested after 4 hours and analyzed on an Automatic Gamma Counter. The specific ⁵¹ Cr dissolution is calculated using the following formula: Percent of specific dissolution = 100 × (test release-spontaneous release) / (maximum release-spontaneous release).

Alternatively, a degranulation assay can be used. For example, K562 cells are subjected to a degranulation assay, followed by determining the percentage of degranulated cells in each lymphocyte subpopulation. Both of these assays are described in WO 2010/110734 (see “3. Evaluation of cell mediated cytotoxicity”). Other in vitro cytotoxicity tests are known in the art.

In one embodiment, the antiviral agent is administered prior to and/or concurrently with and/or after the patient receives NK cell and/or NK-like T cell therapy. It will be appreciated that it is desirable to initiate the administration of the antiviral agent before the reactivation of a human herpes infection is detected or suspected, i. Preferably, the antiviral agent is administered prior to and/or concurrently with the NK cell injection.

By “pre-administer” the inventors include the meaning that the antiviral agent is initially administered before the patient receives the NK cell and/or NK-like T cell therapeutic agent. By “simultaneous administration” we include the meaning that the antiviral agent is initially administered to the patient simultaneously with the NK cell and/or NK-like T cell therapeutic agent. In another embodiment, the antiviral agent is administered after the NK cell and/or NK like T cell therapy. By “administer later” the inventors include the meaning that the antiviral agent is administered first after the patient has received NK cell and/or NK-like T cell therapy.

Preferably, the antiviral agent is administered prior to the onset of the first symptom of viral infection in the patient.

In one embodiment of the invention, the patient is at least one day before the patient receives the NK cell and/or NK-like T cell therapy, such as at least 2 days before the patient receives the NK cell and/or NK-like T cell therapy; Or at least 3 days ago; Or at least 4 days ago; Or at least 5 days ago; Or at least 6 days ago; Or at least 7 days ago; Or at least 8 days ago; Or at least 9 days ago; Or at least 10 days ago; Or at least 20 days ago; Or at least 30 days ago; Alternatively, an antiviral agent is administered at least 1 month before.

In one embodiment, the patient is administered an antiviral agent the day before the patient receives the NK cell and/or NK cell therapeutic agent.

Preferably, the antiviral agent is at least one day after the patient has received NK cell and/or NK-like T cell therapy, such as at least 2 days after the patient has received NK cell and/or NK-like T cell therapy; Or after at least 3 days; Or at least 4 days later.

In one embodiment, the antiviral agent is at least one day after the patient has received an NK cell and/or NK-like T cell therapeutic agent, such as at least two days after the patient has received an NK cell and/or NK-like T cell therapeutic agent; Or after at least 3 days; Or after at least 4 days; Or after at least 5 days; Or after at least 6 days; Or after at least 1 week; Or after at least 2 weeks; Or after at least 3 weeks; Or at least 1 month later.

The dose or amount of the antiviral agent administered to the patient should be a therapeutically effective amount for the intended purpose, ie prophylactic and/or prophylactic measures, and/or an amount effective to kill or inactivate the virus.

In one embodiment, the antiviral agent is in the range of about 1 to about 100 mg/kg body weight daily, preferably in the range of about 2 to 50 mg/kg/day, most preferably in the range of 3 to 20 mg/kg/day. Is administered in a suitable dose of. The desired dose may conveniently be presented as a single dose or in divided doses administered at appropriate intervals, for example as two, three, four or more sub-doses daily. In one embodiment, 500 to 1500 mg of the antiviral agent is administered to the patient within a 24 hour period. In other words, on any given day the patient will receive 500 to 1500 mg of antiviral agent.

The specific therapeutically effective dose for a particular patient will depend on the disorder being treated and the severity of the disorder; The activity of the specific agent(s) used; The age, weight, general health, sex, and diet of the patient; Time of administration; Route of administration; The rate of excretion of the specific agent(s) used; Duration of treatment; It will depend on a variety of factors, including drugs used in combination or coincidence with the particular agent(s) employed and similar factors well known in the medical field. For example, it is within the art to initiate dosing of an agent at a lower level than necessary to achieve the desired therapeutic effect and to gradually increase the dosage until the desired effect is achieved. If desired, the effective daily dose can be divided into multiple doses for administration purposes. Consequently, a single dose composition may contain such amounts or divisors to constitute a daily dose.

Thus, in one embodiment, the antiviral agent is administered to the patient in one or more doses. In one embodiment, the antiviral agent is administered to the patient in 1, 2, 3, 4, 5 or 6 doses. In a preferred embodiment, the antiviral agent is administered to the patient in two doses. In other words, patients receive antiviral drugs twice daily.

Preferably, the antiviral agent is 250 to 1500 mg in a 24 hour period, such as 250 mg in a 24 hour period; Or 300 mg over a 24-hour period; Or 400 mg over a 24-hour period; Or 500 mg over a 24-hour period; Or 600 mg over a 24-hour period; Or 700 mg over a 24-hour period; Or 800 mg over a 24-hour period; Or 900 mg over a 24-hour period; Or 1000 mg over a 24-hour period; Or 1100 mg over a 24-hour period; Or 1200 mg over a 24-hour period; Or 1300 mg over a 24-hour period; Or 1400 mg over a 24-hour period; Alternatively, it is administered to the patient at a dose of 1500 mg over a 24-hour period.

Preferably, the antiviral agent is administered to the patient in two doses of 250 to 750 mg in a 24 hour period, and preferably the antiviral agent is administered to the patient in two doses of 500 mg in the 24 hour period.

It will be appreciated that administration of an antiviral agent can occur as a single event or over the course of treatment. For example, one or more antiviral agents are hourly (e.g., hourly, 2 hourly, 3 hourly, 4 hourly, 5 hourly, 6 hourly, etc.), daily, 2 days, weekly, 2 weeks, or monthly. Can be administered. Certain conditions can extend the prevention from days to weeks. For example, prophylaxis may extend beyond 1, 2, or 3 weeks, or prophylaxis may extend from several days to several weeks.

Thus, in one embodiment, the antiviral agent is at least 1 month, such as at least 2 months; Or at least 3 months; Or at least 4 months; Or at least 5 months; Or at least 6 months; Or administered to the patient for a period of at least 7 months. In a further embodiment, the antiviral agent is administered to the patient for a period of 100 days or less. In a preferred embodiment, the antiviral agent is administered to the patient for a period of up to 7 months, such as 6 months. In a further preferred embodiment, 500 mg of the antiviral agent is administered to the patient twice every 24 hours for a period of 6 months.

It will be understood that in some cases the antiviral agent is administered for a period of more than 7 months, such as 8 or 9 months.

In one embodiment, the patient receives NK cell and/or NK-like T cell therapy after high dose therapy (HDT) and/or autologous stem cell transplantation (ASCT).

By “high-dose therapy (HDT)” we include high-dose chemotherapy, also referred to as “intensive therapy”. In patients with multiple myeloma, HDT is Melphalan (trade name: Alkeran), cyclophosphamide (trade name: Cytoxan), doxorubicin (trade name: Adriamycin), liposomal doxorubicin (trade name: Doxil), and/or panobinostat (tradename). : Farydak) chemotherapy. HDT may include several doses of chemotherapy.

By “autologous stem cell transplantation (ASCT)” we include transplants comprising stem cells obtained from the patient's own blood or bone marrow.

In an alternative embodiment, the stem cell transplant is an allogeneic transplant, wherein the stem cells or bone marrow are obtained from a donor with a matching tissue type (eg, inbred). In an alternative embodiment, the stem cell transplant is syngeneic transplant, wherein the stem cells or bone marrow are obtained from identical twins.

Preferably, stem cell transplantation is autologous (ASCT) (Gertz, MA, & Dingli, D. (2014). How we manage autologous stem cell transplantation for patients with multiple myeloma. Blood, 124(6), 882-890.Accessed March 25, 2018]).

High-dose chemotherapy followed by autologous peripheral blood stem cell transplantation (ASCT) is currently the standard treatment approach for patients under the age of 65 with multiple myeloma.

In a further embodiment, the patient receives NK cell and/or NK-like T cell therapeutic agents 3 to 7 months after ASCT, such as 3 months, or 4 months, or 5 months, or 6 months, or 7 months after ASCT. Receive. Preferably, the patient receives NK cell and/or NK-like T cell therapy 6 months after ASCT.

In one embodiment, the NK cell and/or NK-like T cell therapeutic agent comprises one, or two, or three, or four, or five administrations of NK cells and/or NK-like T cells. Preferably, the patient receives three doses of NK cells and/or NK-like T cells.

In a further embodiment, the NK cells and/or NK-like T cells are 5x10 ⁶ to 100x10 ⁶ cells/kg patient body weight, for example at least 5x10 ⁶ cells/kg patient body weight; Or at least 50x10 ⁶ cells/kg patient body weight; Or at a dose of at least 100×10 ⁶ cells/kg patient body weight. In a preferred embodiment, the patient receives 5x10 ^6, 50x10 ^6, and three times administration of 100x10 ⁶ cells / kg NK cells and / or NK similar increase the capacity of weight or less T-cells in a week interval.

As described above, antiviral agents applicable in the context of the present invention include adhesion to host cells to infect new host cells; Release of viral genes and/or enzymes into the host cell; Replication of viral components using host-cell machinery; Assembly of viral components into complete viral particles; And the release of viral particles at any stage in the life cycle of the virus selected from one or more.

Typical antiviral agents used against herpes virus act by interfering with viral replication, effectively slowing down the rate of replication of the virus, and providing a greater opportunity for the immune response to intervene.

Antiviral agents may be selected from the group comprising substances that act on viral DNA polymerases such as nucleoside analogs after phosphorylation with triphosphate form and phosphonoformic acid and phosphonoacetic acid and analogs thereof.

In some embodiments, the antiviral agent is valomacyclovir stearate (EPB-348), octadecyloxyethyl-cidofovir (ODE-CDV, CMX-001), hexadecyloxypropyl-sidofovir (HDP-CDV). , Abacavir, Adefovir, Amantadine, Amprenavir, Arvidol, Atzanavir, Atrifla, Combivir, Darunavir, Delavirdin, Didanosine, Docosanol, Edoxudin, Efavirens, M Tricitabine, Enfuvirtide, Entecavir, Entry Inhibitor, Antiretrovirus, Pomivirsen, Fosamprenavir, Fusion Inhibitor, Gardacil, Ibacitabine, Imunovir, Idoxuridin, Imiquimod, Indinavir, Inosine , Intergrase inhibitor, interferon type III, interferon type II, interferon type I, interferon, lamivudine, lopinavir, lopinavir, robinid, MK-0518, maraviroc, moroxidin, nelfinavir, nevirapine , Nexavir, nucleoside analogs, oseltamivir, ferramivir, pleconaril, podophylotoxin, protease inhibitor, reverse transcriptase inhibitor, ribavirin, rimantadine, ritonavir, saquinavier, stavudine, synergistic enhancer, Tenofovir, Tenofovir Disproxil, Tipranavier, Trifluridine, Trizivir, Tromantadine, Truvada, Valabiblovir, Vicriviroc, Vidarabine, Viramidine, Zalcitabine, Zanami Beer, zidovudine, A-5021([1'S,2'R)-9[[1'2'-bis(hydroxymethyl)cycloprop-1'-yl]-methyl]guanine]), cyclopro Faber (CPV, ZSM-l-62), 2,4-diamino-6-R43-hydroxy-2 (phosphonomethoxy) propoxy]-pyrimidine (HPMPO-DaPy), N-(4 -Chlorobenzyl)-1-methyl-6-(4-morpholinylmethyl)-4-oxo-1,4-dihydro-3-quinolinecarboxamide (PNU-183792), 2-bromo-5, 6-dichloro-1 -(beta-D-ribofuranosyl)benzimidazole (BDCRB), 1-(beta-L-ribofuranosyl)-2-isopropylamino-5,6-dichlorobenzimidazole (Maribavir , 1263W94), 3-hydroxy-2,2-dimethyl-N[4-{[(5-dimethylamino)-1-naphthyl-sulfonyl}-ami No)phenyl}propamide (BAY 38-4766), 4-(2-amino-4-thiazolyl)phenyl derivative (BILS 179BS), N45-(aminosulfonyl)-4-methyl-1,3-thia Zol-2-yl]-N-methyl-2-{4-(2-pyridinyl)phenyl}acetamide (BAY 57-1293), 2H-3-(4-chlorophenyl)-3,4-dihydro -1,4-benzo-thiazine-2-carbonitrile-1-oxide or 1,1-dioxide and 2-chloro-3-pyridin-3-yl-5,6,7,8-tetrahydroindolizine- It is an antiviral agent selected from the group containing 1-carboxamide (CMV423).

Thus, in one embodiment, the antiviral agent is valacyclovir; Acyclovir; Famcyclovir; And/or a nucleoside analog, such as one selected from the group consisting of pencyclovir, or an active metabolite, prodrug, salt, solvate or hydrate of such a nucleoside analog. Other nucleoside analogs with antiviral activity can be identified by standard methods known in the art. In one embodiment, the antiviral agent is valacyclovir or a prodrug or salt thereof; Acyclovir or a prodrug or salt thereof; Famcyclovir or a prodrug or salt thereof and/or fencyclovir or a prodrug or salt thereof.

It will be appreciated that the antiviral agents of the present invention may comprise a mixture of one or more antiviral agents. In a preferred embodiment, 500 mg of valacyclovir is administered to the patient in two doses within a 24 hour period for a period of 6 months before and during the NK cell and/or NK like T cell therapy.

As used herein, the term “prodrug” refers to a compound that has been transformed in vivo to produce a disclosed compound or a pharmaceutically acceptable form of the compound. In some embodiments, a prodrug is a compound that can be converted by solvolysis under physiological conditions or to a biologically active compound as described herein. Thus, the term “prodrug” refers to a precursor of a pharmaceutically acceptable biologically active compound. The prodrug may be inactive when administered to a patient, but is then converted in vivo to the active compound, for example by hydrolysis (eg, hydrolysis in blood or tissue). In certain cases, the prodrug has improved physical and/or delivery properties compared to the parent compound to which the prodrug is delivered. Prodrugs usually offer the advantage of solubility, tissue compatibility or sustained release in mammalian organisms.

Both acyclovir and pencyclovir are guanosine analogs that inhibit viral replication by acting as a substrate for viral DNA polymerase. Valacyclovir is a prodrug, an esterified version of acyclovir with higher oral bioavailability. Famcyclovir is a prodrug of fencyclovir with improved oral bioavailability. The antiviral agents of Zovirax® (acyclovir), Valtrex® (valacyclovir), Denavir® (phencyclovir) and Famvir® (famcyclovir) are all analogs of acyclic guanosine and are commercially available.

Other nucleoside analogs with antiviral activity are known in the art and can be used in the context of the present invention. For example, bromovinyl deoxyuridine (Brivudin) is a highly potent thymidine nucleoside analog with selective activity against HSV-1 and VZV. It is understood that bicyclic pyrimidine nucleoside analogs (BCNA) have antiviral activity.

Antiviral agents for use in the context of the present invention also include vaccines. Zostavax® is a live, attenuated Varicella zoster vaccine used to prevent zoster-related post-shingles neuralgia (PHN), a long-lasting nerve pain after shingles and shingles. The vaccine can be injected subcutaneously (SC) or intramuscularly (IM). Shingrix® is a non-live, recombinant subunit Varicella zoster vaccine used to prevent shingles (zoster) and reduce the overall incidence of PHN. It combines the antigen, glycoprotein E and the adjuvant system. The vaccine can be injected intramuscularly (IM).

It will be understood by those skilled in the art that the antiviral agent of the present invention may also be used in the form of a pharmaceutically acceptable salt or solvate thereof. Pharmaceutically acceptable salts of antiviral agents include conventional salts formed from pharmaceutically acceptable inorganic or organic acids or bases, and quaternary ammonium salts. More specific examples of suitable acid salts are hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid, perchloric acid, fumaric acid, acetic acid, propionic acid, succinic acid, glycolic acid, formic acid, lactic acid, maleic acid, tartaric acid, citric acid, palmoic acid, malonic acid, hydroxy Maleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, naphthalene-2-sulfonic acid, benzenesulphonic hydroxynaphthoic acid, hydroiodic acid, malic acid, steroic acid, tannic acid, etc. do. Other acids, such as oxalic acid, by themselves are not pharmaceutically acceptable, but may be useful in the preparation of salts useful as intermediates in obtaining the compounds of the present invention and pharmaceutically acceptable salts thereof. More specific examples of suitable base salts are sodium, lithium, potassium, magnesium, aluminum, calcium, zinc, N,N'-dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, N-methylglucamine and Includes procaine salts.

Antiviral agents can be administered by parenteral administration. By parenteral administration we include any means of parenteral administration such as direct injection into the body to bypass the skin and mucous membranes. Common parenteral routes are intramuscular (IM), subcutaneous (SC) and intravenous (IV).

In one embodiment of the present invention, the antiviral agent is administered orally, intravenously, subcutaneously, intravenously and/or intramuscularly.

In one embodiment, Varicella zoster virus (VZV); Herpes simplex virus (HSV); Epstein Barr virus (EBV); And/or a herpes virus selected from the group comprising cytomegalovirus (CMV) is present in the patient. In one embodiment, the herpes virus is human (latent). It will be understood that this patient may be referred to as “serum positive”.

“Serapositive” includes the presence of antibodies or other immune markers in the serum from a patient, which refers to past exposure to a particular organism, antigen or virus. Varicella zoster virus (VZV) prior to the patient receiving treatment comprising natural killer (NK) cells and/or NK-like T cells; Herpes simplex virus (HSV); Epstein Barr virus (EBV); And it will be understood that it may be seropositive to one or more herpes viruses selected from the group comprising cytomegalovirus (CMV). Methods of determining the presence of a latent virus in a patient, such as PCR, such as RT-PCR, are known in the art.

In one embodiment, the herpes virus infection causes shingles.

It will be appreciated that antiviral agents are used to prevent shingles in patients receiving therapeutic agents comprising natural killer (NK) cells and/or NK-like T cells. Severe shingles complications, such as zoster-related post-shingles neuralgia (PHN), multiple shingles, myelitis, ocular herpes, or in patients receiving treatments with antiviral agents comprising natural killer (NK) cells and/or NK-like T cells. It will be understood that it is also used to prevent shingles free.

In a fourth aspect, the present invention provides natural killer (NK) cells and/or NK-like T cells for use in treating malignant diseases in a patient, wherein the use is NK cells and/or NK-like T cells And administering an antiviral agent to the patient receiving the treatment.

In a fifth aspect, the present invention provides the use of natural killer (NK) cells and/or NK-like T cells in the manufacture of a medicament for treating malignant diseases in a patient, wherein the patient is NK cells and/or NK-like T cells. Receive antiviral drugs along with T cell therapy.

In a sixth aspect, the present invention provides a method of treating a malignant disease in a patient, the method comprising administering a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, NK cells And/or administering an antiviral agent to the patient receiving NK-like T cell therapy.

In one embodiment of the invention, the antiviral agent prevents a herpes virus infection in the patient. In one embodiment of the invention, the herpes virus infection comprises reactivation of the herpes virus. It will be appreciated that the herpes virus may include those described above with respect to the first, second and third aspects of the invention.

In one embodiment of the invention, the NK cell and/or NK-like T cell therapeutic agent induces and/or increases herpes virus reactivation. It will be understood that NK cell and/or NK-like T cell therapeutic agents include those described above with respect to the first, second and third aspects of the invention.

In one embodiment of the invention, the patient receives NK cell and/or NK-like T cell therapy after high-dose therapy (HDT) and/or autologous stem cell transplantation.

In one embodiment of the invention, the antiviral agent is as defined above in the context of the first, second and third aspects of the invention.

In a seventh aspect, the invention provides a pharmaceutical composition comprising NK cells and/or NK-like T cells and an antiviral agent as defined above in the context of the first, second and third aspects of the invention. do.

In one embodiment, the pharmaceutical composition further comprises a pharmaceutically acceptable diluent, carrier or excipient. By "pharmaceutically acceptable" it is included that the composition or formulation is sterile and free of pyrogens. Suitable pharmaceutical carriers, diluents and excipients are well known in the pharmaceutical art. The carrier(s) must be "acceptable" in that it is compatible with the inhibitor and is not harmful to its beneficiaries. In general, the carrier will be sterile and pyrogen-free water or saline, although other acceptable carriers may be used.

It will be understood that the pharmaceutical composition comprises a therapeutically effective amount and/or an amount effective to kill or inactivate the virus for its intended purpose, ie prophylactic and/or prophylactic measures.

In one embodiment of the invention, the pharmaceutical composition is for use in preventing a herpes virus infection in a patient, wherein the patient is receiving a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells. , Uses include administering the pharmaceutical composition to a patient receiving NK cell and/or NK-like T cell therapy.

In a further embodiment, the invention provides the use of a pharmaceutical composition in the manufacture of a medicament for preventing herpes virus infection in a patient, wherein the patient comprises natural killer (NK) cells and/or NK-like T cells. And the pharmaceutical composition is administered to a patient receiving NK cell and/or NK-like T cell therapy.

In one embodiment, the invention provides a method of preventing a herpes virus infection in a patient, wherein the patient is receiving a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, the method comprising NK cells and And/or administering the pharmaceutical composition to a patient receiving NK-like T cell therapy.

The following are pharmaceutical preparations and/or compositions according to the invention wherein the active ingredient is an antiviral agent as defined herein.

Although the most suitable route may vary depending on, for example, the condition, age and disorder of the recipient, and the viral infection or disease being treated, the formulation or composition may be administered orally and parenterally (subcutaneous administration, such as injection or depot tablets. And those suitable for intradermal administration, intrasecondary administration, intramuscular administration, including those by depot and intravenous).

In one embodiment, the pharmaceutical composition or formulation of the present invention is suitable for parenteral administration, more particularly for intravenous administration or subcutaneous administration. In a preferred embodiment, the pharmaceutical composition is suitable for intravenous or subcutaneous administration to a patient, for example by injection.

Formulations or compositions suitable for parenteral administration include aqueous and non-aqueous sterile injectable solutions which may contain antioxidants, buffers, bacteriostatic agents and solutes that render the formulation isotonic with the blood of the intended recipient; And aqueous and non-aqueous suspensions which may include suspending agents and thickening agents.

Preferably, the formulation is a unit dose containing a daily dose or unit of the active ingredient, a daily subdose or an appropriate fraction thereof.

The agent or active ingredient is in the form of a pharmaceutical preparation containing the active ingredient, optionally in the form of a non-toxic, organic or inorganic, acid or base, addition salt, orally or any parenteral in a pharmaceutically acceptable dosage form. It can be administered by route. The composition can be administered in various doses depending on the disorder and patient being treated, and the route of administration.

In human treatment, the agent or active ingredient will generally be administered in admixture with a suitable pharmaceutical excipient, diluent or carrier selected with respect to the intended route of administration and standard pharmaceutical practice.

For example, agents or active ingredients may be orally in the form of tablets, capsules, vaginal suppositories, elixirs, solutions or suspensions, which may contain flavorings or coloring agents, for fast-acting, sustained or controlled release applications. , Can be administered buccally or sublingually. The active ingredient can also be administered via intracavernosal injection.

Suitable tablets include excipients such as microcrystalline cellulose, lactose, sodium citrate, calcium carbonate, dibasic calcium phosphate and glycine, disintegrants such as starch (preferably corn, potato or tapioca starch), sodium starch glycolate, croscarmellose. Sodium and certain complex silicates, and granulation binders such as polyvinylpyrrolidone, hydroxypropylmethylcellulose (HPMC), hydroxy-propylcellulose (HPC), sucrose, gelatin and acacia. Additionally, lubricants such as magnesium stearate, stearic acid, glyceryl behenate and talc may be included.

Solid compositions of a similar type can also be used as fillers in gelatin capsules. Preferred excipients in this context include lactose, starch, cellulose, lactose or high molecular weight polyethylene glycol. For aqueous suspensions and/or elixirs, the compounds of the present invention may be combined with various sweeteners or flavors, coloring substances or dyes, emulsifiers and/or suspending agents and diluents such as water, ethanol, propylene glycol and glycerin, and combinations thereof. I can.

The agent or active ingredient may also be administered parenterally, for example intravenously, intraarterial, intraperitoneal, intrathecal, intraventricular, substernal, intracranial, intramuscular or subcutaneous, or by infusion techniques. have. They are best used in the form of sterile aqueous solutions that may contain sufficient salts or other substances such as glucose to render the solution isotonic with blood. The aqueous solution should be suitably buffered if necessary (preferably to a pH of 3 to 9). Preparation of suitable parenteral preparations under sterile conditions is readily accomplished by standard pharmaceutical techniques well known to those skilled in the art.

These formulations may be presented in unit-dose or multi-dose containers, e.g. sealed ampoules and vials, and stored in freeze-dried (lyophilized) conditions requiring only the addition of a sterile liquid carrier, e.g. water for injection, immediately prior to use. Can be. Immediate injection solutions and suspensions can be prepared from sterile powders, granules and tablets of the kind previously described.

For oral and parenteral administration to human patients, the daily dosage level of the agent, antibody or compound will usually be 1 to 1,000 mg per adult (i.e., about 0.015 to 15 mg/kg), single dose or divided dose. Is administered.

Thus, for example, tablets or capsules of the agent or active ingredient may contain 1 mg to 1,000 mg of the agent or active substance, either alone or for two or more administrations at a time, if appropriate. In any case, the physician will determine the actual dosage that will be best suited for the individual patient, which will vary depending on the age, weight and response of the particular patient. The above dosage is an example of the average case. Of course, there may be individual instances where a higher or lower dosage range would be an advantage, and this is within the scope of the present invention.

The agent or active ingredient can also be administered intranasally or by inhalation, and conveniently suitable propellants such as dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoro-ethane, hydrofluoroalkane, Drying by use of, for example, 1,1,1,2-tetrafluoroethane (HFA 134A3 or 1,1,1,2,3,3,3-heptafluoropropane (HFA 227EA3), carbon dioxide or other suitable gas Delivered in the form of a powder inhaler or an aerosol spray tip from a pressurized container, pump, spray or nebulizer In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. , Sprays or nebulizers may contain solutions or suspensions of the active compounds, for example using a mixture of ethanol as a solvent and a propellant, which may further contain a lubricant, for example sorbitan trioleate. Capsules and cartridges for use in the blower (eg, made from gelatin) may be formulated to contain a powder mix of the active ingredient and a suitable powder base, such as lactose or starch, such formulations, for example small cell lung carcinoma , Non-small cell lung carcinoma, pleural pulmonary blastoma or carcinoid tumors, such as solid tumors of the lung.

The aerosol or dry powder formulation is preferably arranged so that each metered dose or “puff” contains at least 1 mg of inhibitor for delivery to a patient. It will be appreciated that the total daily dose by the aerosol will vary from patient to patient and may be administered as a single dose or more generally divided doses over the day.

Alternatively, the agents or active ingredients can be administered in the form of suppositories or pessaries, particularly to treat or target colon, rectal or prostate tumors.

In one embodiment, the agent or active ingredient can be delivered using an injectable sustained release drug delivery system. They are specifically designed to reduce the frequency of injections. An example of such a system is a Nutropin depot encapsulating recombinant human growth hormone (rhGH) in biodegradable microspheres that slowly release rhGH over a prolonged period of time when injected.

The agent or active ingredient may be administered by means of a surgically implanted device that releases the drug directly into the eye, for example, to a site necessary to treat an ocular tumor. This direct application to the disease site achieves effective treatment without significant systemic side effects.

An alternative method for delivery of agents or active ingredients is the heat sensitive Regel injection system. Regel is an injectable liquid below body temperature, while at body temperature it immediately forms a gel reservoir that slowly decays and dissolves with a known safe biodegradable polymer. The active drug is delivered over time as the biopolymer dissolves.

Polypeptide drugs can also be delivered orally. This process uses the natural process for oral absorption of vitamin B ₁₂ from the body to deliver proteins and peptides simultaneously. By using the vitamin B12 absorption system, proteins or peptides can move through the barrier. The complex is synthesized between the drug and the vitamin B ₁₂ analogue, which possesses both a significant affinity for the intrinsic factor (IF) in the vitamin B ₁₂ portion of this complex and a significant bioactivity of the drug portion of this complex. do.

Polynucleotides can be administered as suitable genetic constructs as described below and delivered to patients in which they have been expressed. In general, the polynucleotide in the genetic construct is operably linked to a promoter capable of expressing the compound in the cell. The genetic construct of the present invention can be prepared using a method well known in the art, for example, Sambrook et al. (2001).

The genetic construct for the delivery of polynucleotides may be DNA or RNA, but preferably it is DNA.

Preferably, the genetic construct is adapted for delivery to human cells. Means and methods for introducing genetic constructs into cells are known in the art, and immunoliposomes, liposomes, viral vectors (vaccinia, modified vaccinia, lentivirus, parvovirus, retrovirus, adenovirus and Adeno-associated virus (AAV) vectors), and direct delivery of DNA, for example using gene guns and electrophoresis. In addition, methods of delivering polynucleotides to a target tissue of a patient for treatment are also well known in the art. In an alternative method, a highly efficient nucleic acid delivery system is used that utilizes receptor mediated inclusion to transport DNA macromolecules into cells. This is achieved by conjugating the iron-carrying protein transferrin to a polycation that binds to a nucleic acid. See Cotten et al (1992) Proc. Natl. Acad. Sci. USA 89, 6094-6098] using the endosome-destructive activity of defective adenovirus particles or chemically inactivated adenovirus particles, high-efficiency receptor-mediated DNA constructs or other gene constructs of the present invention Transmission can also be used. It will be appreciated that “naked DNA” and DNA complexed with cationic and neutral lipids may also be useful for introducing the DNA of the invention into the cells of the subject being treated. Nonviral approaches to gene therapy are described in 1995, Human Gene Therapy 6, 1129-1144.

Although it may be useful to use a tissue-specific promoter in a vector encoding a polynucleotide inhibitor for cancer/tumor of a specific tissue, the risk of expression of the active ingredient in a body location other than the cancer/tumor suffers from cancer/tumor. This is not essential as it is expected to be tolerant compared to the therapeutic benefit to the patient. It is desirable to be able to temporarily regulate the expression of the polynucleotide inhibitor in the cell, but this is also not essential.

The agents and/or active ingredients of the invention (ie, antiviral agents) can be lyophilized for storage and reconstituted in a suitable carrier prior to use. Any suitable lyophilization method (eg, spray drying, cake drying) and/or reconstitution techniques may be used. Those skilled in the art will recognize that lyophilization and reconstitution can lead to varying degrees of loss of protein activity and that the level of use needs to be up-regulated to compensate. In one embodiment, the lyophilized (lyophilized) active ingredient, when rehydrated, has its activity (prior to lyophilization) of about 20% or less, or about 25% or less, or about 30% or less, or about 35% or less. , Or less than about 40%, or less than about 45%, or less than about 50%.

It will be understood that the amount of antiviral agent required for use in a preventive measure will vary with the nature of the condition being treated and the age and condition of the patient, and will ultimately depend on the decision of the attending physician. However, in general, the dosages used for adult human treatment will generally range from 0.02 to 5000 mg daily, preferably 100 to 1500 mg daily. The desired dose may conveniently be presented as a single dose or in divided doses administered at appropriate intervals, for example as two, three, four or more sub-doses daily. The formulations according to the invention may contain from 0.1% to 99% of the active ingredient, conveniently from 30% to 95% for tablets and capsules and from 3% to 50% for liquid formulations.

Of course, one of ordinary skill in the art can modify the formulations within the teachings herein to provide a number of formulations for a particular route of administration without destabilizing the compositions of the present invention or impairing their therapeutic activity. It is also well within the ability of those skilled in the art to modify the route of administration and dosage regimen of a particular compound to manage the pharmacokinetics of this compound for maximum benefit effect to the patient.

In the eighth aspect, the present invention

(i) a composition comprising NK cells and/or NK-like T cells, wherein the NK cells and/or NK-like T cells are a composition as defined herein; And

(ii) Provides a kit of parts comprising an antiviral agent as defined herein.

In one embodiment, the kit further comprises a pharmaceutically acceptable diluent, carrier or excipient, such as those described above in the context of a pharmaceutical composition. In a further embodiment, the kit further comprises instructions for administering NK cells and/or NK-like T cells and/or antiviral agents to the patient.

In one embodiment, as defined above in the context of the first, second and third aspects of the invention, the antiviral agent is for use in preventing herpes virus reactivation in a patient.

In a preferred embodiment, as defined above in the context of the first, second and third aspects of the invention, the herpes virus infection comprises reactivation of the herpes virus.

In one embodiment, the NK cell and/or NK-like T cell therapeutic agent induces and/or increases herpes virus reactivation, as defined above in the context of the first, second and third aspects of the invention. .

In one embodiment of the present invention the kit of parts is for use in preventing a herpes virus infection in a patient, wherein the patient receives a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, and uses Comprises administering the components of the kit of parts to a patient receiving NK cell and/or NK-like T cell therapy.

In a further embodiment, the invention provides the use of a kit of parts in the manufacture of a medicament for preventing herpes virus infection in a patient, wherein the patient comprises natural killer (NK) cells and/or NK-like T cells. Upon receiving the therapeutic agent, the components of the parts kit are administered to a patient receiving NK cell and/or NK-like T cell therapy.

In one embodiment, the invention provides a method of preventing a herpes virus infection in a patient, wherein the patient is receiving a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, the method comprising NK cells and And/or administering the components of the parts kit to a patient receiving NK-like T cell therapy.

It will be understood that the features of the fourth, fifth, sixth, seventh and eighth aspects of the present invention may be as described herein in connection with other aspects of the present invention.

All documents cited herein are incorporated herein by reference in their entirety.

Lists or discussions of documents expressly previously published herein should not necessarily be regarded as an admission that the document is part of the state of the art or is of ordinary general knowledge.

The invention will now be described with reference to the following drawings and examples.
Preferred non-limiting embodiments embodying certain aspects of the invention will be described with reference to the following figures:
Figure 1: "CellProtect" safety study
Schematic Chart: Safety Study of “CellProtect”, an autologous ex vivo expanded and activated NK cell product in patients with multiple myeloma (ACP-001).
Figure 2: Clinical efficacy of "CellProtect" treatment
Six patients were injected with three doses of “CellProtect” (indicated by arrows). Two patients had a measurable M component upon infusion. Both of these patients responded with at least a 50% reduction in serum M component (biomarker for tumor burden). This reduction is thought to be clinical "response" (Pt 103) and (Pt 105). After 5 months of infusion, one of these patients relapsed (Pt 105). One patient with a detectable M component showed an improved response after CellProtect injection (Pt 107). The other 3 patients did not show any amount of M component prior to infusion (not shown). None of the patients in complete remission (CR) did not relapse during the course of treatment and clinical follow-up.
Figure 3: "CellProtect" injection and reactivation timeline
Summary of infusion and reactivation timelines in the context of past ASCT in the first 4 patients. "Patient 1" is patient 103, "patient 2" is patient 105, "patient 3" is patient 106, and "patient 4" is patient 107.
Figure 4: Correlation between the time from ASCT to NK cell injection and the time from NK cell injection to the occurrence of HZ
Graph showing the correlation between the time from ASCT to NK cell injection and the time from NK cell injection to the occurrence of HZ. "Patient 3" is patient 103, "patient 4" is patient 105, "patient 5" is patient 106, and "patient 6" is patient 107.
Figure 5: Manufacturing process of "CellProtect" drug substance (DS) and drug product (DP)
Flow chart showing the manufacturing process of CellProtect.

Example 1-Signs of shingles after injection of autologous ex vivo expanded NK cells in patients with multiple myeloma (MM): the need for antiviral prevention

Introduction

The development of new drugs for the treatment of multiple myeloma (MM) has significantly improved survival rates from about 3 years previously to more than 5 years now. However, despite this drastic improvement, the proportion of young patients treated with allogeneic stem cell transplantation (AlloSCT) is a possible exception and cure is never actually achieved. Therefore, a new approach is important.

MM is a malignant neoplasm characterized by clonal proliferation of plasma cells in the bone marrow (BM). This is thought to be impossible to cure due to the persistence of microscopic residual disease despite both novel treatment and intensive treatment (1). The present inventors have previously found that long-term ex vivo expanded and activated autologous NK cells from MM patients provide cytotoxic activity against autologous myeloma cells in vitro, which is superior to short-term activated autologous NK cells ( 2). The inventors have also found that these cells can be used as an effective MM treatment in experimental animals (3).

Recently, the inventors have optimized procedures for NK cell expansion in closed automated bioreactors using clinical grade GMP compliant ingredients (4), completed all preclinical requirements, and completed the first phase I/II clinical trials in humans. To begin with, approval was obtained from the Swedish Drug Licensing Agency (EudraCT: 2010-022330-83) and the Ethics Committee (EPN: 2013/490-32). The expanded cells are fully compliant with the new EU ATMP Directives.

The goal of this stage I/II study of patients previously treated with autologous stem cell transplantation (ASCT) is to first investigate the safety of expanded and activated autologous NK cells, and secondly, monoclonal immunoglobulin, international myeloma research. It is to analyze efficacy parameters such as microscopic residual disease after NK cell therapy in patients responding to the International Myeloma Working Group (IMWG) criteria and responding to ASCT.

Advanced Therapy Medicinal Product (ATMP) CellProtect is a cell suspension based on expanded polyclonal NK cells ex vivo with restored cytotoxic activity. Products are manufactured individually, and the treatment is self-derived.

The first phase I/II, therapeutic exploratory clinical trial in humans by CellProtect in a newly diagnosed patient with multiple myeloma was conducted in 2014 in the Department of Hematology, Karolinska University Hospital, Hudinge. EudraCT 2010-022330-83) (Fig. 1). Key patient inclusion criteria are described in Greipp PR, San Miguel J, Durie BG, et al. (2005), Eligible for, and willing to undergo, high dose chemotherapy and ASCT and Eastern Cooperative Oncology Group (ECOG) performance status 0-2].

The clinical study is an open, single arm, triple ascending dose/patient study to primarily investigate the safety and tolerability of CellProtect in patients with MM after ASCT. The secondary objective was to investigate the effect of CellProtect to reduce response, i.e. to further reduce the M protein in patients who have not achieved complete remission or to reach less microscopic residual disease (MRD) in patients who have achieved complete remission. will be.

result

Six patients completed the study including evaluation and follow-up for 6 months after the last injection (FIG. 1 ). Interim analysis of clinical outcomes according to the protocol is completed with the option of closing the study, with safety data determined to be sufficient, alternatively keeping it open and including an additional 6 patients.

Research patients will initially donate blood for the manufacture of CellProtect after being included in the study. The starting material is collected through regular blood donations and transferred to a manufacturing facility where the final product CellProtect is manufactured. CellProtect is stored in cryopreservation at -150°C at the dose adjusted to the patient until requested by the principal investigator's location. The available bag(s) is delivered to the clinic in liquid nitrogen by the manufacturer, where it is thawed just before injection.

The shelf life of the active substance is based on the ongoing stability monitoring program from CellProtect's validation badge. Based on this data, it is concluded that the CellProtect drug product (DP) is stable at -150°C for at least 48 months. It is expected that additional data will be collected and the shelf life of the final product may be extended. The stability in use of the active substance was determined to be at least 60 minutes.

Patients were treated according to current clinical practice with 3 to 4 cycles Cyber-D (cyclophosphamide, bortezomib, dexamethasone) as induced following blood donation followed by high-dose treatment (melphalan 200 mg/m2) And stem cell injection.

Study treatment is initiated when the patient recovers from ASCT but within 6 months from ASCT. 5x10 ⁶ patients per week; 50x10 ⁶ pcs.; And 3 CellProtect injections at an elevated dose of 100×10 ⁶ cells/kg body weight or less. There is a dose escalation within each patient. CellProtect treatment was evaluated during the 6 month follow-up period after the last injection. The effectiveness of treatment during this follow-up period was assessed in greater depth at Study Visit 7, approximately one month after the third CellProtect injection. At Visit 7, separate blood samples and bone marrow material were collected for exploratory analysis of specific immunogenic responses to CellProtect treatment at this point. This sample will be analyzed by a method designed to specifically evaluate and explore the mechanism of action of the CellProtec Investigative Medical Product (IMP).

Clinical results

Clinical research status

Six (6) patients successfully completed according to the protocol CellProtect treatment, and three infusions according to the protocol requested the accumulated number of activated NK cells. These 6 patients were evaluated at Visit 7, followed by a 6-month follow-up.

Patient population studied

Patients are included in clinical trials at diagnosis, and patient demographic data are listed in Table 1. Five patients (103, 105, 106, 107, 111) had IgG myeloma and one patient (110) had IgA myeloma. The response state after ASCT and before NK cell injection was very good partial response (VGPR) in 3 patients (103, 105, 107) and complete response (CR) in 3 patients (106, 110, 111). The starting material for the production of CellProtect is collected by blood donation at the first study visit prior to initiation of any MM treatment.

Response to high-dose chemotherapy and ASCT

The second study visit is a confirmation visit after ASCT and before CellProtect injection. The purpose of this visit is to establish the patient's classification according to MM Response Criteria, Table 2, and to collect information about the patient's physical condition to ensure that the patient is still eligible and well enough to continue the study treatment. Bone marrow samples are taken at this visit, informed consent must be provided from the patient, and constitute a baseline BM sampling for evaluation of CellProtect treatment. The patient's status transfer at Visit 3 serves as the baseline for most other assessments.

Preliminary clinical results

Patient 103 was classified as a very good partial response (VGPR) to ASCT treatment and received three full doses of CellProtect infusions 6 to 8 weeks after implantation. M component serum levels decreased from 8 g/L before CellProtect injection to 1 g/L after injection (>80% reduction), which was low throughout the study period.

Patient 105 was classified as a very good partial response (VGPR) to ASCT treatment and received 3 full doses of CellProtect infusions 12 to 14 weeks after implantation. The decrease in component M serum levels was measured from 5 g/L to 2 g/L (approximately 60%) after CellProtect injection and remained low for 4 months. The patient relapsed 5 months after CellProtect injection. Patient 105 disagrees and therefore no bone marrow sampling was taken at Visit 2.

Patient 106 was classified as a complete response to ASCT treatment (CR) and received three doses of CellProtect at 15 to 17 weeks post implantation. The third dose decreased due to lack of IMP. The patient was in complete remission (CR) throughout clinical follow-up.

Patient 107 was initially classified as a very good partial response (VGPR) and later confirmed complete remission to ASCT treatment. Patient 107 had detectable levels of component M at the time of CellProtect injection, and three full doses of CellProtect were injected 15 to 30 weeks after implantation. The third and highest dose of CellProtect was delayed due to the activation of herpes zoster and signs of shingles in the patient. Patient 107 had an improved response after CellProtect injection. A decrease in M component serum levels was observed from 1 g/L to 0 g/L. The free light chain (FLC) quota (736 at screening) in this patient decreased from 1.6 to 0.8 during and after CellProtect infusion. The patient was in complete remission (CR) throughout clinical follow-up.

Patient 110 was classified as a complete response to ASCT treatment (CR) and received three doses of CellProtect at 14 to 16 weeks post implantation. The third dose decreased due to lack of IMP. The patient was in complete remission (CR) throughout clinical follow-up.

Patient 111 was classified as complete response to ASCT treatment (CR) and received three full doses of CellProtect infusions 17 to 19 weeks after transplantation. The patient was in complete remission (CR) throughout clinical follow-up.

efficacy

Signs of the clinical efficacy of CellProtect treatment have been monitored to this day by measuring serum immunoglobulin levels, an established biomarker for disease (FIG. 2 ). The decrease in serum immunoglobulin levels was measured after administration of CellProtect in all (3) patients with remaining measurable disease and was in stable partial response (VGPR) or (VGPR) after ASCT. Three of the six (6) patients (3) were in complete remission (CR) and did not have measurable levels of serum immunoglobulins in CellProtect treatment. The increase in immunoglobulin levels was measured to date in one of six (6) patients treated with CellProtect (FIG. 2 ).

Interestingly, activation of herpes zoster and signs of shingles in this clinical trial were observed in the first four patients (103, 105, 106 and 107) dosed with CellProtect. Patients 110 and 111 were treated with high-dose precautions prior to CellProtect injection to prevent virus activation. This side effect of CellProtect products further confirms the biological activity of the cell preparation in vivo.

safety

There were no serious side effects. However, the first 4 patients (103, 105, 106 and 107) developed herpes zoster (HZ, shingles) 18 to 32 weeks after ASCT and 3 to 25 weeks after the first NK cell injection ( Fig. 3). These patients received valacyclovir 250 mg x 2 daily for 14 weeks after ASCT as a prevention of virus reactivation according to clinical practice. Thereafter, the drug was canceled, and accordingly, the time at which HZ occurred after NK cell injection was not included.

Since the onset of HZ is clearly associated with NK cell infusion in the first 4 patients, the last 2 patients (110 and 111) were treated with 500 mg of high dose valacyclovir twice daily for 6 months after infusion. These two patients did not show any signs of HZ activation within 12 months after prophylactic treatment.

The infusion and reactivation timelines in the context of previous ASCT of the first 4 patients are summarized in Figure 3.

Review

Our results show that self-derived expanded and activated NK cells are safe, but induce HZ reactivation if antiviral drugs are not used for prophylaxis.

It seems clear that NK cell injection is the cause of the occurrence of HZ. HZ is always visible after NK cell injection, and as shown in FIG. 4, there is an inverse correlation between the time from ASCT to NK cell injection and the time from NK cell injection to the occurrence of HZ.

A possible mechanism for HZ development is the induction of an immunological cascade response upon adoptive activated NK cell translocation. NK cells activated upon adoptive cell transfer are designed to attack reservoir cells for HZV, eventually causing viral reactivation due to induced stress.

Considering the time of HZV onset associated with the time of ASCT and NK cell injection, respectively, ASCT in particular will not be the cause of HZV activation. In addition, recent reports show only 1.0% to 14.4% of HZV infection in ASCT-treated MM patients without antiviral prophylaxis (8, 9), and as in our study, 4 out of 4 consecutive patients (100%). Is not likely to develop shingles from ASCT.

Our conclusion is that although NK cell based immunotherapy is feasible in MM, it should always be combined with prophylactic antiviral therapy.

Materials and methods

Manufacturing of CellProtect

The CellProtect drug product is a cell suspension based on NK cells expanded ex vivo from patients with MM. Treatment is autologous.

The protocol for making CellProtect is described below and shown in Figure 5.

Peripheral blood from patients with MM is collected via blood donation. According to the blood donation method, one-unit (about 450 ml) of whole blood is collected in a sterile polyvinylchloride (PVC) plastic carrying bag. (Terumo or Fenwal blood collection vessel). The donated blood is stored at room temperature (15-25° C.), and the manufacturing process starts within 6 hours.

Lymphocytes are separated by a density-based gradient using Ficoll-Paque. After the final washing step with PBS, the cells are counted and adjusted to 0.5 to 1.0 × 10 ⁶ cells/mL in 800 to 1000 mL of cell culture medium supplemented with the following materials: 500 IU/mL of interleukin 2 (IL -2, cytokine activating NK cells), 10 ng/mL Orthoclone OKT3 (muromonam-CD3, CD-3 antibody that stimulates the growth of T cells), 5% (v/v) human serum (stimulating growth ) And 0.1% (v/v) Pluronic F68 (a surfactant that reduces foaming). The cells are seeded in the bioreactor and culture begins.

Peripheral blood lymphocytes are expanded using a closed Wave bioreactor system (System 2/10 GE Healthcare) that controls the temperature to 37°C and 5% CO2. Cells are grown in a disposable cellbag 2 L (culture volume 1 L). The cell contacting surface is an ethylene vinyl acetate (EVA)/low density polyethylene copolymer. The outer layer is made from a proprietary composite that provides exceptional strength and extremely low gas permeability.

When the cell density reaches 3 × 10 ⁶ cells/mL, perfusion is initiated by feeding the culture to the cell medium supplemented with the above-described substances except that it does not have OKT3. Perfusion is controlled in a shot of 50 mL of cell culture medium containing 500 IU/mL of IL-2, 5% human serum and 0.1% Pluronic F68. Cell concentration determines the volume of the shot.

The NK cell expansion phase begins when perfusion begins and lasts for 14 to 16 days.

Cell number and viability are monitored on a regular basis during the culture period. Flow cytometry analysis is performed to determine the percentage of NK cells. Activated NK cells are analyzed by expression of a surrogate marker for cytotoxic CD107a after triggering by the K562 cell line. Sterility, endotoxin and mycoplasma tests are also performed on samples taken after expansion during the manufacture of CellProtect DP.

Specifications of CellProtect Investigative Medicine Products (IMP)

Tables 4 and 5 below describe the specifications of CellProtect products.

Composition of clinical badges

The composition of the CellProtect clinical badge is shown in Table 6.

Example 1 References:

1. Alici E, Bjorkstrand B, Treschow A, Aints A, Smith CI, Gahrton G, et al. Long-term follow-up of gene-marked CD34+ cells after autologous stem cell transplantation for multiple myeloma. Cancer Gene Ther. 2007;14(3):227-32.

2. Alici E, Sutlu T, Bjorkstrand B, Gilljam M, Stellan B, Nahi H, et al. Autologous antitumor activity by NK cells expanded from myeloma patients using GMP-compliant components. Blood. 2008;111(6):3155-62.

3. Alici E, Konstantinidis KV, Sutlu T, Aints A, Gahrton G, Ljunggren HG, et al. Anti-myeloma activity of endogenous and adoptively transferred activated natural killer cells in experimental multiple myeloma model. Exp Hematol. 2007;35(12):1839-46.

4. Sutlu T, Stellan B, Gilljam M, Quezada HC, Nahi H, Gahrton G, et al. Clinical-grade, large-scale, feeder-free expansion of highly active human natural killer cells for adoptive immunotherapy using an automated bioreactor. Cytotherapy. 2010;12(8):1044-55.

5. Backstrom E, Chambers BJ, Ho EL, Naidenko OV, Mariotti R, Fremont DH, et al. Natural killer cell-mediated lysis of dorsal root ganglia neurons via RAE1/NKG2D interactions. European journal of immunology. 2003;33(1):92-100.

6. Backstrom E, Chambers BJ, Kristensson K, Ljunggren HG. Direct NK cell-mediated lysis of syngenic dorsal root ganglia neurons in vitro. Journal of immunology. 2000;165(9):4895-900.

7. Hickey WF, Ueno K, Hiserodt JC, Schmidt RE. Exogenously-induced, natural killer cell-mediated neuronal killing: a novel pathogenetic mechanism. The Journal of experimental medicine. 1992;176(3):811-7.

8. Park H, Youk J, Kim HR, Koh Y, Kwon JH, Yoon SS, et al. Infectious complications in multiple myeloma receiving autologous stem cell transplantation in the past 10 years. International journal of hematology. 2017;106(6):801-10.

9. Park S, Jung CW, Jang JH, Kim SJ, Kim WS, Kim K. Incidence of infection according to intravenous immunoglobulin use in autologous hematopoietic stem cell transplant recipients with multiple myeloma. Transpl Infect Dis. 2015;17(5):679-87.

Example 2: Pharmaceutical formulation

The following examples illustrate the pharmaceutical formulations according to the present invention in which the active ingredient is an antiviral agent.

Example A: Tablet

Active ingredient 100 mg

Lactose 200 mg

Starch 50 mg

Polyvinylpyrrolidone 5 mg

Magnesium stearate 4 mg

359 mg

Tablets are prepared by wet granulation of the above ingredients and then compression.

Example B: Ophthalmic Solution

Active ingredient 0.5 g

Sodium chloride, analytical grade 0.9 g

Thiomersal 0.001 g

Purified water 100 ml

pH adjusted 7.5

Example C: Tablet formulation

Formulations A and B below are prepared by wet granulation of the ingredients with a povidone solution, followed by addition of magnesium stearate and compression.

Formulation A

mg/tablet mg/tablet

(a) active ingredient 250 250

(b) Lactose B.P. 210 26

(c) Povidone B.P. 15 9

(d) sodium starch glycolate 20 12

(e) magnesium stearate 5 3

500 300

Formulation B

mg/tablet mg/tablet

(a) active ingredient 250 250

(b) lactose 150 -

(c) Avicel PH 101® 60 26

(d) Povidone B.P. 15 9

(e) sodium starch glycolate 20 12

(f) magnesium stearate 5 3

500 300

Formulation C

mg/tablet

Active ingredient 100

Lactose 200

Starch 50

Povidone 5

Magnesium stearate 4

359

Formulations D and E below are prepared by directly compressing the mixed ingredients. The lactose used in Formulation E is in direct compressed form.

Formulation D

mg/capsule

Active ingredient 250

Pregelatinized starch starch NF15 150

400

Formulation E

mg/capsule

Active ingredient 250

Lactose 150

Avicel ® 100

500

Formulation F (controlled release formulation)

This formulation is prepared by wet granulation of the component (below) with a povidone solution, followed by addition of magnesium stearate and compression.

mg/tablet

(a) active ingredient 500

(b) hydroxypropylmethylcellulose 112

(Methocel K4M Premium)®

(c) lactose B.P. 53

(d) Povidone B.P.C. 28

(e) magnesium stearate 7

700

Drug release occurs over a period of about 6 to 8 hours and is complete after 12 hours.

Example D: Capsule formulation

Formulation A

The capsule formulation is prepared by mixing the ingredients of Formulation D in Example C above, and filling it into a two-part hard gelatin capsule. Formulation B (below) is prepared in a similar manner.

Formulation B

mg/capsule

(a) active ingredient 250

(b) Lactose B.P. 143

(c) sodium starch glycolate 25

(d) magnesium stearate 2

420

Formulation C

mg/capsule

(a) active ingredient 250

(b) Macrogol 4000 BP 350

600

Capsules are prepared by melting Macrogol 4000 BP, dispersing the active ingredient in the melt, and filling the melt into two part hard gelatin capsules.

Formulation D

mg/capsule

Active ingredient 250

lecithin 100

Arakis Oil 100

450

Capsules are prepared by dispersing the active ingredient in lecithin and arachis oil, and filling the dispersion with soft elastic gelatin capsules.

Formulation E (controlled release capsule)

The following controlled release capsule formulation is prepared by extruding components a, b and c using an extruder, followed by spheroidizing and drying the extrudate. Thereafter, the dried pellets are coated with a release-controlling film (d), and filled with a two-part hard gelatin capsule.

mg/capsule

(a) active ingredient 250

(b) microcrystalline cellulose 125

(c) lactose BP 125

(d) ethyl cellulose 13

513

Example E: Formulation for Injection

Active ingredient 0.200 g

10 ml sterile pyrogen-free phosphate buffer (pH 7.0)

The active ingredient is dissolved in most of the phosphate buffer (35 to 40° C.) to make it volume, filtered through a sterile micropore filter into a 10 ml sterile amber glass vial (type 1), and sealed with a sterile stopper and overseal. do.

Example F: Intramuscular injection

Active ingredient 0.20 g

Benzyl alcohol 0.10 g

Glucofurol 75® 1.45 g

Water for injection 3.00 ml

The active ingredient is dissolved in glycolfurol. Then, benzyl alcohol is added and dissolved, and water is added up to 3 ml. Thereafter, the mixture is filtered through a sterile microporous filter and sealed with a 3 ml sterile glass vial (Type 1).

Example G: Syrup suspension

Active ingredient 0.2500 g

Sorbitol solution 1.5000 g

Glycerol 2.0000 g

Dispersible cellulose 0.0750 g

Sodium benzoate 0.0050 g

Flavoring, peach flavor 17.42.3169 0.0125 ml

Purified water appropriate amount 5.0000 ml

Sodium benzoate is dissolved in a portion of purified water, and a sorbitol solution is added. Add active ingredient and disperse. The thickener (dispersible cellulose) is dispersed in glycerol. The two dispersions are mixed and brought to the required volume with purified water. Additional thickening is achieved as needed by additional suspension shear.

Example H: Suppositories

mg/suppository

Active ingredient (63 μm)* 250

Hard Fat, BP (Witepsol H15-Dynamit Nobel) 1770

2020

* The active ingredient is used as a powder in which at least 90% of the particles have a diameter of 63 μm or less.

One-fifth of Witepsol H15 is melted in a steam jacketed pan at 45°C maximum. The active ingredient is sieved through a 200 μm sieve and added to the molten base while mixing using a silverson equipped with a cutting head until a smooth dispersion is achieved. While maintaining the mixture at 45° C., the remaining Witepsol H15 is added to the suspension and stirred to ensure a homogeneous mixture. The entire suspension is passed through a 250 μm stainless steel screen and allowed to cool to 40° C. with continuous stirring. At a temperature of 38° C. to 40° C., 2.02 g of the mixture is charged into a suitable plastic mold. Allow the suppository to cool to room temperature.

Example I: Pessary

mg/pessary

Active ingredient 250

Anhydride dextrose 380

Potato starch 363

Magnesium stearate 7

1000

The above ingredients are directly mixed and the resulting mixture is directly compressed to prepare a pessary.

Claims (40)

As an antiviral agent for use in preventing herpes virus reactivation in a patient, the patient receives a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, and uses NK cells and/or Or, an antiviral agent comprising administering an antiviral agent to a patient receiving NK-like T cell therapy. As the use of an antiviral agent in the manufacture of a medicament for preventing herpes virus reactivation in a patient, the patient receives a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, and the antiviral agent is NK cells and/or Or to a patient receiving NK-like T cell therapy. As a method of preventing herpes virus reactivation in a patient, the patient is receiving a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, and the method is receiving NK cells and/or NK-like T cell therapy. A method comprising administering an antiviral agent to the patient. An antiviral agent for the use of claim 1, the use of claim 2, or the method of claim 3, wherein the NK cell and/or NK-like T cell therapeutic agent induces and/or increases herpes virus reactivation. Or how. The antiviral agent, use or method for use according to any one of the preceding claims, wherein the patient has a malignant disease. The antiviral agent, use or method for use according to any one of the preceding claims, wherein the NK cell and/or NK-like T cell therapeutic agent is for use in the treatment of a malignant disease. The antiviral agent, use or method for use according to claim 5 or 6, wherein the malignant disease is hematologic cancer. The antiviral agent, use or method for use according to claim 7, wherein the hematologic cancer is selected from the group consisting of myeloma, lymphoma, leukemia and/or chronic myeloproliferative diseases. The antiviral agent, use or method for use according to any one of the preceding claims, wherein the NK cells have the phenotype CD3 ^- CD56 ⁺ and/or the NK like T cells have the phenotype CD3 ⁺ CD56 ⁺ . The antiviral agent, use or method for use according to any one of the preceding claims, wherein the NK cells and/or NK-like T cells are expanded ex vivo. The antiviral agent for use according to any one of the preceding claims, wherein the antiviral agent is administered before and/or at the same time as and/or after receiving the NK cell and/or NK-like T cell therapeutic agent by the patient. Way. The method of any one of the preceding claims, wherein the patient is at least one day before the patient receives NK cell and/or NK-like T cell therapy, such as at least 2 days before the patient receives NK cell and/or NK-like T cell therapy; Or at least 3 days ago; Or at least 4 days ago; Or at least 5 days ago; Or at least 6 days ago; Or at least 7 days ago; Or at least 8 days ago; Or at least 9 days ago; Or at least 10 days ago; Or at least 20 days ago; Or at least 30 days ago; Or an antiviral agent, use or method for use, wherein the antiviral agent is administered at least 1 month before. The method according to any one of the preceding claims, wherein the antiviral agent is at least one day after the patient has received NK cell and/or NK-like T cell therapy, such as at least 2 days after the patient has received NK cell and/or NK-like T cell therapy. ; Or after at least 3 days; Or an antiviral agent, use or method for use, administered after at least 4 days. The antiviral agent, use or method for use according to any one of the preceding claims, wherein 500 to 1500 mg of the antiviral agent is administered to the patient within a 24 hour period. The antiviral agent, use or method for use according to any one of the preceding claims, wherein the antiviral agent is administered to the patient in one or more doses. According to any one of the preceding claims, the antiviral agent is administered to the patient in a dose of 250 to 750 mg 2 in a 24-hour period, preferably the antiviral agent is administered to the patient in a double dose of 500 mg in a 24-hour period. An antiviral agent, use or method for use, administered. The method according to claim 1, wherein the antiviral agent is at least 1 month, such as at least 2 months; Or at least 3 months; Or at least 4 months; Or at least 5 months; Or at least 6 months; Or an antiviral agent, use or method for use, administered to a patient for a period of at least 7 months. The method of any one of the preceding claims, wherein the patient is receiving NK cells and/or NK-like T cell therapy after high dose therapy (HDT) and/or autologous stem cell transplantation (ASCT). , Antiviral agent for use, use or method. The method of claim 18, wherein the patient is 3 to 7 months after ASCT, such as 3 months after ASCT, or 4 months after ASCT, or 5 months after ASCT, or 6 months after ASCT, or 7 months after ASCT. An antiviral agent, use or method for use, receiving an NK cell and/or NK-like T cell therapeutic agent. The method according to any one of the preceding claims, wherein the NK cell and/or NK-like T cell therapeutic agent is one, or two, or three, or four, or five or more times of NK cells and/or NK-like T cells. Antiviral agents, uses or methods for use, including administration. The method of any one of the preceding claims, wherein the NK cells and/or NK-like T cells are at least 5x10 ⁶ cells/kg patient body weight; Or at least 50x10 ⁶ cells/kg patient body weight; Or an antiviral agent, use or method for use, administered at a dosage of at least 100×10 ⁶ cells/kg patient body weight. The method of any one of the preceding claims, wherein the antiviral agent is valacyclovir; Acyclovir; Famcyclovir; And/or a nucleoside analog as selected from the group consisting of pencyclovir or an active metabolite, prodrug, salt, solvate or hydrate of such a nucleoside analog. An antiviral agent, use or method for use. The antiviral agent, use or method for use according to any one of the preceding claims, wherein the antiviral agent is administered orally, intravenously, subcutaneously and/or intramuscularly. According to any one of the preceding claims, Varicella zoster virus (VZV: varicella zoster virus); Herpes simplex virus (HSV); Epstein Barr Virus (EBV); And/or a herpes virus selected from the group consisting of cytomegalovirus (CMV) is present in a patient, an antiviral agent, use or method for use. The antiviral agent, use or method for use according to any one of the preceding claims, wherein the herpes virus reactivation causes shingles in a patient. The antiviral agent, use or method for use according to any one of the preceding claims, wherein the patient is not immunodepleted. As natural killer (NK) cells and/or NK-like T cells for use in treating malignant diseases in a patient, the use of which is to administer an antiviral agent to a patient receiving NK cells and/or NK-like T cell therapy. Natural killer (NK) cells and/or NK-like T cells for use, comprising the steps of. Use of natural killer (NK) cells and/or NK-like T cells in the manufacture of a medicament for the treatment of malignant diseases in a patient, wherein the patient is administered an antiviral agent together with NK cells and/or NK-like T cell therapeutic agents, Usage. A method of treating a malignant disease in a patient, comprising administering a therapeutic agent comprising natural killer (NK) cells and/or NK-like T cells, and receiving NK cells and/or NK-like T cell therapy to a patient The method further comprising administering an antiviral agent. Natural killer (NK) cells for use according to any one of claims 27 to 29, wherein the malignant disease is a hematologic cancer such as selected from the group consisting of myeloma, lymphoma, leukemia and/or chronic myeloproliferative disease. And/or NK-like T cells, uses or methods. 31. The use or method of any one of claims 27 to 30, wherein the antiviral agent prevents herpes virus reactivation in a patient, for use in natural killer (NK) cells and/or NK-like T cells. The NK cell and/or NK-like T cell therapeutic agent for use according to any one of claims 27 to 31, which induces and/or increases herpes virus reactivation, and/or NK Similar T cells, uses or methods. 33. Natural killer (NK) cells and/or NK-like T cells for use, use or method according to claim 31 or 32, wherein the herpes virus reactivation causes shingles in the patient. 33.The natural killing (NK) cell of any one of claims 27-32, wherein the patient is receiving NK cells and/or NK-like T cell therapeutics following high-dose therapy (HDT) and/or autologous stem cell transplantation. ) Cells and/or NK-like T cells, uses or methods. The method according to any one of claims 27 to 34, wherein the antiviral agent is a natural killer (NK) cell and/or NK-like T cell, method or use for use, as defined in any one of the preceding claims. . The method of any one of claims 27-35, wherein the NK cell and/or NK-like T cell therapeutic agent is a natural killer (NK) cell for use and/or as defined in any one of the preceding claims. NK-like T cells, methods or uses. A pharmaceutical composition comprising natural killer (NK) cells and/or NK-like T cells as defined in any of the preceding clauses, and an antiviral agent as defined in any of the preceding clauses, Pharmaceutical composition. As a kit of parts,
(i) a composition comprising natural killer (NK) cells and/or NK-like T cells, wherein the NK cells and/or NK-like T cells are as defined in any one of the preceding paragraphs; And
(ii) A kit of parts comprising an antiviral agent as defined in any of the preceding paragraphs. The pharmaceutical composition of claim 37 or the kit of claim 38, further comprising a pharmaceutically acceptable diluent, carrier, or excipient. An antiviral agent, use, method, pharmaceutical composition or kit of parts for use, substantially as described herein in connection with the appended claims and examples.

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